Injectable biodegradable polymeric particles (usually microspheres) represent an exciting approach to control the release of vaccine antigens to reduce the number of doses in the immunization schedule and optimize the desired immune response via selective targeting of antigen to antigen presenting cells. After the first couple of decades of their study, much progress has been made towards the clinical use of antigen-loaded microspheres. Poly(lactide-co-glycolic acids) (PLGAs) have been studied most commonly for this purpose because of their proven safety record and established use in marketed products for controlled delivery of several peptide drugs. PLGA microspheres have many desirable features relative to standard aluminum-based adjuvants, including the microspheres' ability to induce cell-mediated immunity, a necessary requirement for emergent vaccines against HIV and cancer. PLGA microparticles have displayed unprecedented versatility and safety to accomplish release of one or multiple antigens of varying physical-chemical characteristics and immunologic requirements, and have now met numerous critical benchmarks in development of long-lasting immunity after a single injected dose.
Chances are that for every important protein that has undergone pharmaceutical development, the polymer-controlled release option, at the very least, has been considered seriously and, in many cases, aggressively pursued. Unfortunately, successful controlled release of proteins has been a daunting task and, until recently, there has been significant doubt whether a significant number of therapeutic proteins could be slowly and completely released in a native state from the biodegradable polymer-of-choice for general biomedical applications, copolymers from lactic and glycolic acids. Thus, the most significant obstacle in the development of controlled-release injectable depots for proteins has emerged as the instability of the protein during encapsulation and release in vivo.
Hydrophilic macromolecules, like proteins, cannot diffuse through the hydrophobic polymer phase, like through PLGA. The release of encapsulated protein drugs from PLGA requires at some point the diffusion of the macromolecules through water-filled pores and channels. Protein release from PLGA microspheres typically follows a tri-phasic behavior. First, protein on the surface or having access to the surface of microspheres (i.e., in open pores) is released rapidly, which is the source of the initial burst release. Then, there is a lag time because protein cannot diffuse through the polymer phase. The continuous protein release will not occur until polymer erosion begins, which will produce more pores and channels and consequently let protein in previously isolated pores release out.
Most theoretical frameworks to date regarding protein release have neglected the dynamics of polymer microstructure; namely the kinetics of pore opening and closing.
Current methods for encapsulating molecules, such as biomacromolecules in biodegradable polymers involves harsh processing conditions, such as organic solvents, excess heat, mixing and so forth, which can denature and/or destabilize proteins and other biomacromolecules. Additionally, drying and micronization of biomacromolecules, which often occurs prior to encapsulations may further destabilize the biomacromolecules.
Accordingly, a need exists for new methods of encapsulating biomacromolecules in pore-containing polymers, such as PLGA. The method should be able to be performed without need for organic solvent or other harsh processing conditions during encapsulation, which can denature proteins or destabilize other biomacromolecules. There should also be no need for micronization of the protein or poly(nucleic acid) before encapsulation, which can destabilize both biomacromolecules. A need exists for a method of encapsulation wherein there is no need for drying of the biomacromolecule, which can also destabilize this species.
A need further exists for a method wherein the polymer microspheres could be acceptably terminally sterilized (e.g., by gamma irradiation) before encapsulation with small losses in polymer molecular weight, wherein the sterile protein (or biomacromolecule)-containing solution and sterile microspheres could be placed in a syringe and microencapsulation could be performed at the point-of-care, or sterile protein solution could be added to sterile microspheres as typically done with a diluent being added to typical dry microspheres that already contain protein.
There also exists a need for new methods of encapsulation which are less expensive to carry out than conventional methods, which may be a principal factor in the slow development of more controlled release injectable depots. Furthermore the new method should be applicable to tissue engineering scaffolds, presumably, and any type of biomaterial (or any other polymer encapsulation system, e.g., agricultural) that requires the need to encapsulate molecules that do not strongly partition into the polymer phase, but in pores (typically aqueous as in biomaterials) of the polymer.